5.1 A bioconcentration test is conducted to obtain information concerning the ability of an aquatic species to accumulate a test material directly from water. This guide provides guidance for designing bioconcentration tests on the properties of the test material so that each material is tested in a cost-effective manner.

5.2 Because steady-state is usually approached from the low side and the definition of apparent steady-state is based on a statistical hypothesis test, the apparent steady-state BCF will usually be lower than the steady-state BCF. With the variation and sample sizes commonly used in bioconcentration tests, the actual steady-state BCF will usually be no more than twice the apparent BCF.

5.3 When both are determined in the same test, the projected steady-state BCF will usually be higher than the apparent steady-state BCF because the models used to calculate the projected BCF assume that the BCF steadily increases until infinite time.

5.4 The BCFs and rates and extents of uptake and depuration will depend on temperature, water quality, the species and its size, physiological condition, age, and other factors ( 1 ) . ³ Although organisms are fed during tests, uptake by means of sorption onto food is probably negligible during tests.

5.5 Results of bioconcentration tests are used to predict concentrations likely to occur in aquatic organisms in field situations as a result of exposure under comparable conditions, except that mobile organisms might avoid exposure when possible. Under the experimental conditions, particulate matter is deliberately minimized compared to natural water systems. Exposure conditions for the tests may therefore not be comparable for an organic chemical that has a high octanol-water partition coefficient or for an inorganic chemical that sorbs substantially onto particulate matter. The amount of the test substance in solution is thereby reduced in both cases, and therefore the material is less available to many organisms. However, sorption might increase bioaccumulation by aquatic species that ingest particulate matter ( 2 ) , or food may be a more important source of residues in fish than water per se for stable neutral organic chemicals that have a Log K _ow between 4 and 6 ( 3 ) .

5.6 Results of bioconcentration tests can be used to compare the propensity of different materials to be accumulated. Nonionizable organic chemicals can also be ranked for bioconcentration using correlations that have been reported between steady-state BCFs and physicalchemical properties, such as the octanolwater partition coefficient and solubility in water ( 4 ). However, when such predictions are impossible, exceed the demonstrated limits of the correlation, or might be otherwise questionable ( 1 , 5 ), a bioconcentration test may be necessary.

5.7 Results of bioconcentration tests can also be used to compare the abilities of different species to accumulate materials. At steady-state the concentration of a nonionizable organic chemical in individual organisms, and in various tissues within an organism, will probably be related to the concentration of lipids in the organisms and tissues ( 6 ).

5.8 Results of bioconcentration tests might be an important consideration when assessing hazard (see Guide E1023 ) or deriving water-quality criteria because consumer animals might be adversely affected by ingesting aquatic organisms that contain toxic materials. However, assessment of hazard to consumer organisms must take into account not only the quantity of material accumulated in tissues of aquatic organisms, but also the toxicity of the material to the consumer. Further, humans eat only certain portions of most aquatic organisms, whereas other predators often consume additional tissues.

5.9 Bioconcentration tests might be useful for studying structureactivity relationships between test materials, biological availability, metabolism of materials in aquatic organisms, and effects of various environmental factors on results of such tests.

5.10 Uptake and depuration rate constants might be useful for predicting environmental fate using compartmental models ( 7 ).

1.1 This guide describes procedures for obtaining laboratory data concerning bioconcentration of a test material added to dilution waterbut not to foodby freshwater and saltwater fishes and saltwater bivalve mollusks using the flow-through technique. These procedures also should be useful for conducting bioconcentration tests with other aquatic species, although modifications might be necessary.

1.2 Other modifications of these procedures might be justified by special needs or circumstances. Although using appropriate procedures is more important than following prescribed procedures, the results of tests conducted using unusual procedures are not likely to be comparable to those of many other tests. The comparison of results obtained using modified and unmodified versions of these procedures might provide useful information concerning new concepts and procedures for conducting bioconcentration tests.

1.3 These procedures are applicable to all chemicals that can be measured accurately at the necessary concentrations in water and in appropriate tissues. Bioconcentration tests are usually conducted on individual chemicals but can be conducted on mixtures if appropriate measurements can be made. Some techniques described in this guide were developed for tests on non-ionizable organic chemicals (see 11.1.2.1 ) and might not apply to ionizable or inorganic chemicals.

1.4 Results of bioconcentration tests should usually be reported in terms of apparent steady-state and projected steady-state bioconcentration factors (BCFs) and uptake and depuration rate constants. Results should be reported in terms of whole body for fishes and in terms of total soft tissue for bivalve mollusks. For fishes and scallops consumed by humans, some results should also be reported in terms of the edible portion, especially if ingestion of the test material by humans is a major concern. For tests on organic and organometallic chemicals, the percent lipids of the tissue should be reported.

1.5 This guide is arranged as follows:

1.6 The values stated in SI units are to be regarded as standard. No other units of measurement are included in this standard.

1.7 This standard does not purport to address all of the safety concerns, if any, associated with its use. It is the responsibility of the user of this standard to establish appropriate safety and health practices and determine the applicability of regulatory limitations prior to use. Specific precautionary statements are given in Section 7 .